Substitution of strictly conserved Y111 in catalase-peroxidases: Impact of remote interdomain contacts on active site structure and catalytic performance.

Catalase-peroxidase function is strictly dependent on a gene-duplicated C-terminal domain. This domain no longer has a functioning active site, but from 25 to 30A away it is essential for preventing the coordination of an active site base (His106) to the heme. The mechanisms by which this distant structure supports active site function have not yet been elucidated. Tyr111 is a strictly conserved member of an interdomain H-bonding network that supports the loop connecting the N-terminal B (bearing His106) and C helices. Spectroscopic evaluation of the Tyr111Ala variant of KatG showed a substantial increase in hexa-coordinate low-spin heme, giving it the appearance of a transition between the wild type (primarily high-spin) and the N-terminal domain alone (pure low-spin). Concomitant with the spectral changes was decreased activity compared to the wild type enzyme, suggesting that Tyr111 does have a role in preventing His106 coordination. Substitution of Tyr111 diminishes catalase activity more substantially than peroxidase activity. Such an effect cannot be explained by His106 coordination alone, suggesting that these interdomain interactions may help tune the catalase-peroxidase active site for bifunctionality.